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用DNA对应物替换锤头状核酶的非催化茎环区域,只会增加米氏常数(KM),而不会影响催化步骤(催化常数kcat):一种提高底物特异性的方法。

Substitution of non-catalytic stem and loop regions of hammerhead ribozyme with DNA counterparts only increases KM without sacrificing the catalytic step (kcat): a way to improve substrate-specificity.

作者信息

Shimayama T, Sawata S, Komiyama M, Takagi Y, Tanaka Y, Wada A, Sugimoto N, Rossi J J, Nishikawa F, Nishikawa S

机构信息

Hitachi Chemical Co., Tsukuba Science City, Japan.

出版信息

Nucleic Acids Symp Ser. 1992(27):17-8.

PMID:1283905
Abstract

In elucidating structure-function relationships and stabilizing ribozymes in vivo, several chimeric RNA/DNA ribozymes and substrates were chemically synthesized. Measurements of kinetic parameters revealed that the maximally deoxyribonucleotide-substituted ribozyme (DRDRD32) gained the highest catalytic activity reaching the kcat value of > 10 min-1, the highest value ever reported for hammerhead-type ribozymes. Since these chimeric ribozymes are more stable than the wild-type all-RNA ribozymes in vivo and they also possess higher substrate-specificity, they are considered to be better candidates for antiviral therapeutic agents.

摘要

在阐明体内结构-功能关系以及稳定核酶的过程中,化学合成了几种嵌合RNA/DNA核酶和底物。动力学参数测量结果表明,最大程度脱氧核苷酸取代的核酶(DRDRD32)获得了最高的催化活性,其催化常数(kcat)值>10 min-1,这是锤头型核酶所报道的最高值。由于这些嵌合核酶在体内比野生型全RNA核酶更稳定,并且它们还具有更高的底物特异性,因此它们被认为是抗病毒治疗剂的更好候选者。

相似文献

1
Substitution of non-catalytic stem and loop regions of hammerhead ribozyme with DNA counterparts only increases KM without sacrificing the catalytic step (kcat): a way to improve substrate-specificity.用DNA对应物替换锤头状核酶的非催化茎环区域,只会增加米氏常数(KM),而不会影响催化步骤(催化常数kcat):一种提高底物特异性的方法。
Nucleic Acids Symp Ser. 1992(27):17-8.
2
Nuclease-resistant chimeric ribozymes containing deoxyribonucleotides and phosphorothioate linkages.含有脱氧核糖核苷酸和硫代磷酸酯键的耐核酸酶嵌合核酶。
Nucleic Acids Res. 1993 Jun 11;21(11):2605-11. doi: 10.1093/nar/21.11.2605.
3
Acquisition of novel catalytic activity by the M1 RNA ribozyme: the cost of molecular adaptation.M1 RNA核酶获得新的催化活性:分子适应的代价。
J Mol Biol. 1999 Oct 1;292(4):931-44. doi: 10.1006/jmbi.1999.3098.
4
The effect of universal fluorinated nucleobases on the catalytic activity of ribozymes.通用氟化核碱基对核酶催化活性的影响。
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Comparative single-turnover kinetic analyses of trans-cleaving hammerhead ribozymes with naturally derived non-conserved sequence motifs.对具有天然衍生的非保守序列基序的反式切割锤头状核酶进行的比较单轮动力学分析。
FEBS Lett. 2005 Mar 14;579(7):1619-24. doi: 10.1016/j.febslet.2005.02.016.
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Importance of specific purine-pyrimidine amino and hydroxyl groups for efficient cleavage by a hammerhead ribozyme.特定嘌呤 - 嘧啶氨基和羟基对锤头状核酶高效切割的重要性。
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Enhancement of the cleavage rates of DNA-armed hammerhead ribozymes by various divalent metal ions.多种二价金属离子对DNA武装的锤头状核酶切割速率的增强作用。
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Isolation of active ribozymes from an RNA pool of random sequences using an anchored substrate RNA.使用锚定底物RNA从随机序列的RNA文库中分离活性核酶。
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Construction of new ribozymes requiring short regulator oligonucleotides as a cofactor.需要短调节寡核苷酸作为辅助因子的新型核酶的构建。
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Effects of deoxyribonucleotide substitutions in the substrate strand on hammerhead ribozyme-catalyzed reactions.底物链中脱氧核糖核苷酸取代对锤头状核酶催化反应的影响。
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引用本文的文献

1
Effects of helical structures formed by the binding arms of DNAzymes and their substrates on catalytic activity.脱氧核酶的结合臂与其底物形成的螺旋结构对催化活性的影响。
Nucleic Acids Res. 1998 Jul 15;26(14):3385-91. doi: 10.1093/nar/26.14.3385.
2
Nuclease-resistant chimeric ribozymes containing deoxyribonucleotides and phosphorothioate linkages.含有脱氧核糖核苷酸和硫代磷酸酯键的耐核酸酶嵌合核酶。
Nucleic Acids Res. 1993 Jun 11;21(11):2605-11. doi: 10.1093/nar/21.11.2605.
3
Enhancement of the cleavage rates of DNA-armed hammerhead ribozymes by various divalent metal ions.
多种二价金属离子对DNA武装的锤头状核酶切割速率的增强作用。
Nucleic Acids Res. 1993 Dec 11;21(24):5656-60. doi: 10.1093/nar/21.24.5656.